This project involves the molecular genetic analysis of a set of interacting regulatory mechanisms in bacteriophage P22. Most of the genes of phage P22 are subject to negative control by the P22 c2 repressor (product of gene c2). The P22 ant gene codes for an antirepressor protein, a novel regulatory protein which represents a higher order of complexity in mechanisms of regulation of gene expression, since it works by interacting with and inhibiting the c2 repressor protein. Synthesis of antirepressor is in turn under negative control by two repressors, the products of P22 genes arc and mnt. The nature and mechanism of action of the arc and mnt repressors will be investigated. The arc and mnt gene products will be identified by SDS-polyacrylamide gel electrophoresis by comparing the proteins synthesized in cells infected with P22 wild type, arc-am mutants, and mnt-am mutants. To demonstrate the mechanism of action of these repressors, the rate of synthesis of ant gene mRNA and antirepressor protein will be measured in the presence and absence of the mnt and arc gene products. Experiments are also planned to investigate how the mnt and arc genes are expressed and regulated characterizing mutations and conditions which affect synthesis of the mnt and arc proteins. P22 arc- phage not only overproduce antirepressor, but show other alterations in regulation of many P22 genes. Our current hypothesis to explain this observation is that P22 antirepressor, when produced in vast quantities, interferes with other P22 regulatory proteins. We are investigating this idea by comparing similar alterations in regulation of P22 gene expression.